RESUMO
The family of proglucagon peptides Includes glucagon and glucagon-like peptide 1 (GLP-1), two unique peptides derived from the same prohormone. Despite numerous similarities between the peptides, these have long been viewed as having opposing actions on metabolism. GLP-1 is described as a postprandial hormone that stimulates anabolic actions via insulin, while glucagon is viewed as a fasting hormone that drives catabolic actions to maintain euglycemia. Here, we revisit a classic article in Diabetes that first established that glucagon and GLP-1 have more in common than previously appreciated, including actions at the same receptor. Furthermore, we discuss how the impact of this observation has guided research decades later that has reshaped the view of how proglucagon hormones regulate metabolism.
Assuntos
Glucagon , Insulina , Proglucagon , Peptídeo 1 Semelhante ao Glucagon , JejumRESUMO
During mouse pregnancy placental lactogens stimulate prolactin receptors on pancreatic islet beta cells to induce expression of the tryptophan hydroxylase Tph1, resulting in the synthesis and secretion of serotonin. Presently, the functional relevance of this phenomenon is unclear. One hypothesis is that serotonin-induced activation of 5-HT2B receptors on beta cells stimulates beta cell proliferation during pregnancy. We tested this hypothesis via three different mouse models: (i) total Tph1KO mice, (ii) 129P2/OlaHsd mice, which are incompetent to upregulate islet Tph1 during pregnancy, whereas Tph1 is normally expressed in the intestine, mammary glands, and placenta, and (iii) Htr2b-deficient mice. We observed normal pregnancy-induced levels of beta cell proliferation in total Tph1KO mice, 129P2/OlaHsd mice, and in Htr2b-/- mice. The three studied mouse models indicate that islet serotonin production and its signaling via 5-HT2B receptors are not required for the wave of beta cell proliferation that occurs during normal mouse pregnancy.
Assuntos
Células Secretoras de Insulina , Ilhotas Pancreáticas , Feminino , Animais , Gravidez , Camundongos , Serotonina/metabolismo , Placenta/metabolismo , Ilhotas Pancreáticas/metabolismo , Células Secretoras de Insulina/metabolismo , Proliferação de Células , Triptofano Hidroxilase/genética , Triptofano Hidroxilase/metabolismoRESUMO
During vascular development, endothelial cAMP-dependent protein kinase A (PKA) regulates angiogenesis by controlling the number of tip cells, and PKA inhibition leads to excessive angiogenesis. Whether this role of endothelial PKA is restricted to embryonic and neonatal development or is also required for vascular homeostasis later on is unknown. Here, we show that perinatal (postnatal days P1-P3) of later (P28-P32) inhibition of endothelial PKA using dominant-negative PKA expressed under the control of endothelial-specific Cdh5-CreERT2 recombinase (dnPKAiEC mice) leads to severe subcutaneous edema, hypoalbuminemia, hypoglycemia and premature death. These changes were accompanied by the local hypersprouting of blood vessels in fat pads and the secondary enlargement of subcutaneous lymphatic vessels. Most noticeably, endothelial PKA inhibition caused a dramatic disorganization of the liver vasculature. Hepatic changes correlated with decreased gluconeogenesis, while liver albumin production seems to be unaffected and hypoalbuminemia is rather a result of increased leakage into the interstitium. Interestingly, the expression of dnPKA only in lymphatics using Prox1-CreERT2 produced no phenotype. Likewise, the mosaic expression in only endothelial subpopulations using Vegfr3-CreERT2 was insufficient to induce edema or hypoglycemia. Increased expression of the tip cell marker ESM1 indicated that the inhibition of PKA induced an angiogenic response in the liver, although tissue derived pro- and anti-angiogenic factors were unchanged. These data indicate that endothelial PKA is a gatekeeper of endothelial cell activation not only in development but also in adult homeostasis, preventing the aberrant reactivation of the angiogenic program.
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Vasos Sanguíneos , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico , Células Endoteliais , Fígado , Albuminas , Animais , Vasos Sanguíneos/metabolismo , Vasos Sanguíneos/fisiologia , AMP Cíclico , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/metabolismo , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/fisiologia , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/fisiologia , Células Endoteliais/metabolismo , Células Endoteliais/fisiologia , Homeostase , Hipoalbuminemia , Hipoglicemia , Fígado/metabolismo , Fígado/fisiologia , Camundongos , RecombinasesRESUMO
BACKGROUND: Different types of proteins diverge at vastly different rates. Moreover, the same type of protein has been observed to evolve with different rates in different phylogenetic lineages. In the present study we measured the rates of protein evolution in Eutheria (placental mammals) and Metatheria (marsupials) on a genome-wide basis and we propose that the gene position in the genome landscape has an important influence on the rate of protein divergence. RESULTS: We analyzed a protein-encoding gene set (n = 15,727) common to 16 mammals (12 Eutheria and 4 Metatheria). Using sliding windows that averaged regional effects of protein divergence we constructed landscapes in which strong and lineage-specific regional effects were seen on the molecular clock rate of protein divergence. Within each lineage, the relatively high rates were preferentially found in subtelomeric chromosomal regions. Such regions were observed to contain important and well-studied loci for fetal growth, uterine function and the generation of diversity in the adaptive repertoire of immunoglobulins. CONCLUSIONS: A genome landscape approach visualizes lineage-specific regional differences between Eutherian and Metatherian rates of protein evolution. This phenomenon of chromosomal position is a new element that explains at least part of the lineage-specific effects and differences between proteins on the molecular clock rates.
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Eutérios , Placenta , Animais , Eutérios/genética , Feminino , Genoma/genética , Mamíferos/genética , Filogenia , GravidezRESUMO
FURIN is a proprotein convertase (PC) responsible for proteolytic activation of a wide array of precursor proteins within the secretory pathway. It maps to the PRC1 locus, a type 2 diabetes susceptibility locus, but its specific role in pancreatic ß-cells is largely unknown. The aim of this study was to determine the role of FURIN in glucose homeostasis. We show that FURIN is highly expressed in human islets, whereas PCs that potentially could provide redundancy are expressed at considerably lower levels. ß-cell-specific Furin knockout (ßFurKO) mice are glucose intolerant as a result of smaller islets with lower insulin content and abnormal dense-core secretory granule morphology. mRNA expression analysis and differential proteomics on ßFurKO islets revealed activation of activating transcription factor 4 (ATF4), which was mediated by mammalian target of rapamycin C1 (mTORC1). ßFurKO cells show impaired cleavage or shedding of vacuolar-type ATPase (V-ATPase) subunits Ac45 and prorenin receptor, respectively, and impaired lysosomal acidification. Blocking V-ATPase pharmacologically in ß-cells increased mTORC1 activity, suggesting involvement of the V-ATPase proton pump in the phenotype. Taken together, these results suggest a model of mTORC1-ATF4 hyperactivation and impaired lysosomal acidification in ß-cells lacking Furin, causing ß-cell dysfunction.
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Fator 4 Ativador da Transcrição/metabolismo , Furina/metabolismo , Células Secretoras de Insulina/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Animais , Diabetes Mellitus Tipo 2/metabolismo , Furina/genética , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Transdução de Sinais/fisiologiaRESUMO
BACKGROUND: Circulating donor-specific anti-HLA antibodies (HLA-DSAs) are often absent in serum of kidney allograft recipients whose biopsy specimens demonstrate histology of antibody-mediated rejection (ABMR). It is unclear whether cases involving ABMR histology without detectable HLA-DSAs represent a distinct clinical and molecular phenotype. METHODS: In this multicenter cohort study, we integrated allograft microarray analysis with extensive clinical and histologic phenotyping from 224 kidney transplant recipients between 2011 and 2017. We used the term ABMR histology for biopsy specimens that fulfill the first two Banff 2017 criteria for ABMR, irrespective of HLA-DSA status. RESULTS: Of 224 biopsy specimens, 56 had ABMR histology; 26 of these (46.4%) lacked detectable serum HLA-DSAs. Biopsy specimens with ABMR histology showed overexpression of transcripts mostly related to IFNγ-induced pathways and activation of natural killer cells and endothelial cells. HLA-DSA-positive and HLA-DSA-negative biopsy specimens with ABMR histology displayed similar upregulation of pathways and enrichment of infiltrating leukocytes. Transcriptional heterogeneity observed in biopsy specimens with ABMR histology was not associated with HLA-DSA status but was caused by concomitant T cell-mediated rejection. Compared with cases lacking ABMR histology, those with ABMR histology and HLA-DSA had higher allograft failure risk (hazard ratio [HR], 7.24; 95% confidence interval [95% CI], 3.04 to 17.20) than cases without HLA-DSA (HR, 2.33; 95% CI, 0.85 to 6.33), despite the absence of transcriptional differences. CONCLUSIONS: ABMR histology corresponds to a robust intragraft transcriptional signature, irrespective of HLA-DSA status. Outcome after ABMR histology is not solely determined by the histomolecular presentation but is predicted by the underlying etiologic factor. It is important to consider this heterogeneity in further research and in treatment decisions for patients with ABMR histology.
Assuntos
Rejeição de Enxerto/etiologia , Antígenos HLA/imunologia , Isoanticorpos/sangue , Transplante de Rim/efeitos adversos , Transcrição Gênica , Adulto , Idoso , Feminino , Rejeição de Enxerto/patologia , Sobrevivência de Enxerto , Humanos , Rim/metabolismo , Rim/patologia , Masculino , Pessoa de Meia-Idade , Doadores de Tecidos , Transplante HomólogoRESUMO
BACKGROUND: Antibody-mediated rejection, a leading cause of renal allograft graft failure, is diagnosed by histological assessment of invasive allograft biopsies. Accurate non-invasive biomarkers are not available. METHODS: In the multicentre, prospective BIOMARGIN study, blood samples were prospectively collected at time of renal allograft biopsies between June 2011 and August 2016 and analyzed in three phases. The discovery and derivation phases of the study (Nâ¯=â¯117 and Nâ¯=â¯183 respectively) followed a case-control design and included whole genome transcriptomics and targeted mRNA expression analysis to construct and lock a multigene model. The primary end point was the diagnostic accuracy of the locked multigene assay for antibody-mediated rejection in a third validation cohort of serially collected blood samples (Nâ¯=â¯387). This trial is registered with ClinicalTrials.gov, number NCT02832661. FINDINGS: We identified and locked an 8-gene assay (CXCL10, FCGR1A, FCGR1B, GBP1, GBP4, IL15, KLRC1, TIMP1) in blood samples from the discovery and derivation phases for discrimination between cases with (Nâ¯=â¯49) and without (Nâ¯=â¯134) antibody-mediated rejection. In the validation cohort, this 8-gene assay discriminated between cases with (Nâ¯=â¯41) and without antibody-mediated rejection (Nâ¯=â¯346) with good diagnostic accuracy (ROC AUC 79·9%; 95% CI 72·6 to 87·2, pâ¯<â¯0·0001). The diagnostic accuracy of the 8-gene assay was retained both at time of stable graft function and of graft dysfunction, within the first year and also later after transplantation. The 8-gene assay is correlated with microvascular inflammation and transplant glomerulopathy, but not with the histological lesions of T-cell mediated rejection. INTERPRETATION: We identified and validated a novel 8-gene expression assay that can be used for non-invasive diagnosis of antibody-mediated rejection. FUNDING: The Seventh Framework Programme (FP7) of the European Commission.
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Anticorpos/imunologia , Biomarcadores , Ácidos Nucleicos Livres , Rejeição de Enxerto/diagnóstico , Rejeição de Enxerto/etiologia , Transplante de Rim/efeitos adversos , RNA Mensageiro/genética , Adulto , Feminino , Rejeição de Enxerto/sangue , Humanos , Biópsia Líquida , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , RNA Mensageiro/sangue , Curva ROC , Reprodutibilidade dos Testes , Transplante HomólogoRESUMO
OBJECTIVE: Cytochrome c oxidase (COX) deficiency is a major mitochondrial respiratory chain defect that has vast genetic and phenotypic heterogeneity. This study aims to identify novel causative genes of COX deficiency with only striated muscle-specific symptoms. METHODS: Whole exome sequencing was performed in 2 unrelated individuals who were diagnosed with congenital myopathy and presented COX deficiency in muscle pathology. We assessed the COX6A2 variants using measurements of enzymatic activities and assembly of mitochondrial respiratory chain complexes in the samples from the patients and knockout mice. RESULTS: Both patients presented muscle weakness and hypotonia in 4 limbs along with facial muscle weakness. One patient had cardiomyopathy. Neither patient exhibited involvement from other organs. Whole exome sequencing identified biallelic missense variants in COX6A2, which is expressed only in the skeletal muscle and heart. The variants detected were homozygous c.117C > A (p.Ser39Arg) and compound heterozygous c.117C > A (p.Ser39Arg) and c.127T > C (p.Cys43Arg). We found specific reductions in complex IV activities in the skeletal muscle of both individuals. Assembly of complex IV and its supercomplex formation were impaired in the muscle. INTERPRETATION: This study indicates that biallelic variants in COX6A2 cause a striated muscle-specific form of COX deficiency. ANN NEUROL 2019;86:193-202.
Assuntos
Deficiência de Citocromo-c Oxidase/diagnóstico por imagem , Deficiência de Citocromo-c Oxidase/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Variação Genética/genética , Proteínas Musculares/genética , Músculo Esquelético/patologia , Adolescente , Sequência de Aminoácidos , Animais , Evolução Fatal , Células HEK293 , Células HeLa , Humanos , Lactente , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , LinhagemRESUMO
OBJECTIVES: Peroxisomes play a crucial role in lipid and reactive oxygen species metabolism, but their importance for pancreatic ß-cell functioning is presently unknown. To examine the contribution of peroxisomal metabolism to ß-cell homeostasis in mice, we inactivated PEX5, the import receptor for peroxisomal matrix proteins, in an inducible and ß-cell restricted manner (Rip-Pex5-/- mice). METHODS: After tamoxifen-induced recombination of the Pex5 gene at the age of 6 weeks, mice were fed either normal chow or a high-fat diet for 12 weeks and were subsequently phenotyped. RESULTS: Increased levels of very long chain fatty acids and reduced levels of plasmalogens in islets confirmed impairment of peroxisomal fatty acid oxidation and ether lipid synthesis, respectively. The Rip-Pex5-/- mice fed on either diet exhibited glucose intolerance associated with impaired insulin secretion. Ultrastructural and biochemical analysis revealed a decrease in the density of mature insulin granules and total pancreatic insulin content, which was further accompanied by mitochondrial disruptions, reduced complex I activity and massive vacuole overload in ß-cells. RNAseq analysis suggested that cell death pathways were affected in islets from HFD-fed Rip-Pex5-/- mice. Consistent with this change we observed increased ß-cell apoptosis in islets and a decrease in ß-cell mass. CONCLUSIONS: Our data indicate that normal peroxisome metabolism in ß-cells is crucial to preserve their structure and function.
Assuntos
Células Secretoras de Insulina/metabolismo , Peroxissomos/metabolismo , Animais , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Receptor 1 de Sinal de Orientação para Peroxissomos/deficiência , Receptor 1 de Sinal de Orientação para Peroxissomos/metabolismoRESUMO
BACKGROUND AND AIMS: Early treatment of Crohn's disease [CD] is required in order to optimize patient outcomes. To this end, we need to gain a better understanding of the molecular changes at the onset of CD. METHODS: As a model for the earliest mucosal CD lesions, we study post-operative recurrent CD [Rutgeerts score ≥ i2b]. We are the first to analyse gene and microRNA [miRNA] expression profiles in ileal biopsies from these patients, and compare them with those of newly diagnosed [≤18 months] and late-stage [>10 years after diagnosis] CD patients. RESULTS: Except for one gene [WNT5A], there are no differential genes in CD patients without post-operative recurrence [i0], showing that previous disease did not influence gene expression in the neoterminal ileum, and that this model can be used to study early mucosal CD lesions. Gene expression and co-expression network dysregulation is more pronounced in newly diagnosed and late-stage CD than in post-operative recurrent CD, with most important modules associated with [a]granulocyte adhesion/diapedesis, and cholesterol biosynthesis. In contrast, we found a role for snoRNAs/miRNAs in recurrent CD, highlighting the potential importance of regulatory RNAs in early disease stages. Immunohistochemistry confirmed the expression of key dysregulated genes in damaged/regenerating epithelium and immune cells in recurrent CD. CONCLUSIONS: Aside from regulatory RNAs, there are no clear gene signatures separating post-operative recurrent, newly diagnosed, and late-stage CD. The relative contribution of dysregulated genes and networks differs, and suggests that surgery may reset the disease at the mucosal site, and therefore post-operative recurrent CD might be a good model a good model to study to study early mucosal CD lesions.
Assuntos
Doença de Crohn/genética , Perfilação da Expressão Gênica , MicroRNAs/metabolismo , Adulto , Idoso , Bélgica , Biópsia , Doença de Crohn/cirurgia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , RecidivaRESUMO
Despite partial elucidation of the pathophysiology of antibody-mediated rejection (ABMR) after kidney transplantation, it remains largely unclear which of the involved immune cell types determine disease activity and outcome. We used microarray transcriptomic data from a case-control study (n=95) to identify genes that are differentially expressed in ABMR. Given the co-occurrence of ABMR and T-cell-mediated rejection (TCMR), we built a bioinformatics pipeline to distinguish ABMR-specific mRNA markers. Differential expression of 503 unique genes was identified in ABMR, with significant enrichment of natural killer (NK) cell pathways. CIBERSORT (Cell type Identification By Estimating Relative Subsets Of known RNA Transcripts) deconvolution analysis was performed to elucidate the corresponding cell subtypes and showed increased NK cell infiltration in ABMR in comparison to TCMR and normal biopsies. Other leukocyte types (including monocytes/macrophages, CD4 and CD8 T cells, and dendritic cells) were increased in rejection, but could not discriminate ABMR from TCMR. Deconvolution-based estimation of NK cell infiltration was validated using computerized morphometry, and specifically associated with glomerulitis and peritubular capillaritis. In an external data set of kidney transplant biopsies, activated NK cell infiltration best predicted graft failure amongst all immune cell subtypes and even outperformed a histologic diagnosis of acute rejection. These data suggest that NK cells play a central role in the pathophysiology of ABMR and graft failure after kidney transplantation.
Assuntos
Anticorpos/imunologia , Rejeição de Enxerto/diagnóstico , Falência Renal Crônica/cirurgia , Transplante de Rim/efeitos adversos , Células Matadoras Naturais/imunologia , Adulto , Idoso , Aloenxertos/citologia , Aloenxertos/imunologia , Aloenxertos/patologia , Biomarcadores/análise , Biópsia , Estudos de Casos e Controles , Biologia Computacional , Conjuntos de Dados como Assunto , Feminino , Perfilação da Expressão Gênica , Rejeição de Enxerto/genética , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/patologia , Humanos , Rim/citologia , Rim/imunologia , Rim/patologia , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Valor Preditivo dos Testes , Prognóstico , Resultado do Tratamento , Adulto JovemRESUMO
Vitamin D and all its metabolites are bound to a specific vitamin D binding protein, DBP. This protein was originally first discovered by its worldwide polymorphism and called Group-specific Component (GC). We now know that DBP and GC are the same protein and appeared early in the evolution of vertebrates. DBP is genetically the oldest member of the albuminoid family (including albumin, α-fetoprotein and afamin, all involved in transport of fatty acids or hormones). DBP has a single binding site for all vitamin D metabolites and has a high affinity for 25OHD and 1,25(OH)2D, thereby creating a large pool of circulating 25OHD, which prevents rapid vitamin D deficiency. DBP of higher vertebrates (not amphibians or reptiles) binds with very high affinity actin, thereby preventing the formation of polymeric actin fibrils in the circulation after tissue damage. Megalin is a cargo receptor and is together with cubilin needed to reabsorb DBP or the DBP-25OHD complex, thereby preventing the urinary loss of these proteins and 25OHD. The total concentrations of 25OHD and 1,25(OH)2D in DBP null mice or humans are extremely low but calcium and bone homeostasis remain normal. This is the strongest argument for claiming that the "free hormone hypothesis" also applies to the vitamin D hormone, 1,25(OH)2D. DBP also transports fatty acids, and can play a role in the immune system. DBP is genetically very polymorphic with three frequent alleles (DBP/GC 1f, 1s, and 2) but in total more than 120 different variants but its health consequences, if any, are not understood. A standardization of DBP assays is essential to further explore the role of DBP in physiology and diseases.
RESUMO
Malaria reduces host fitness and survival by pathogen-mediated damage and inflammation. Disease tolerance mechanisms counter these negative effects without decreasing pathogen load. Here, we demonstrate that in four different mouse models of malaria, adrenal hormones confer disease tolerance and protect against early death, independently of parasitemia. Surprisingly, adrenalectomy differentially affects malaria-induced inflammation by increasing circulating cytokines and inflammation in the brain but not in the liver or lung. Furthermore, without affecting the transcription of hepatic gluconeogenic enzymes, adrenalectomy causes exhaustion of hepatic glycogen and insulin-independent lethal hypoglycemia upon infection. This hypoglycemia is not prevented by glucose administration or TNF-α neutralization. In contrast, treatment with a synthetic glucocorticoid (dexamethasone) prevents the hypoglycemia, lowers cerebral cytokine expression and increases survival rates. Overall, we conclude that in malaria, adrenal hormones do not protect against lung and liver inflammation. Instead, they prevent excessive systemic and brain inflammation and severe hypoglycemia, thereby contributing to tolerance.
Assuntos
Glândulas Suprarrenais/metabolismo , Encéfalo/imunologia , Citocinas/imunologia , Hormônios/imunologia , Hipoglicemia/imunologia , Fígado/imunologia , Pulmão/imunologia , Malária/imunologia , Glândulas Suprarrenais/imunologia , Adrenalectomia , Animais , Glicemia/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Corticosterona/imunologia , Corticosterona/metabolismo , Citocinas/efeitos dos fármacos , Dexametasona/farmacologia , Modelos Animais de Doenças , Epinefrina/imunologia , Epinefrina/metabolismo , Glucocorticoides/imunologia , Glucocorticoides/farmacologia , Glicogênio/metabolismo , Hidrocortisona/imunologia , Hidrocortisona/metabolismo , Inflamação , Fígado/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Camundongos , Mineralocorticoides/imunologia , Mineralocorticoides/metabolismo , Norepinefrina/imunologia , Norepinefrina/metabolismo , Plasmodium berghei , Plasmodium chabaudi , Taxa de SobrevidaRESUMO
The minigene encoding human growth hormone (hGH) has been incorporated into over 300 transgenic mouse lines to improve transgene expression. However, unexpected and functional hGH expression can drastically alter physiology. We list here the mouse lines in which ectopic hGH has been confirmed, and we provide a wiki for lines awaiting analysis.
Assuntos
Artefatos , Hormônio do Crescimento Humano/genética , Transgenes/genética , Animais , Humanos , Camundongos , Camundongos TransgênicosRESUMO
OBJECTIVE: Lymphocyte recruitment to the inflamed gut is increased in UC. Inhibition of this cell trafficking by vedolizumab (VDZ) was successful in inducing and maintaining remission and in induction of endoscopic mucosal healing. There are no data on histological healing with VDZ. We studied histological changes following VDZ therapy and compared gene expression in patients with UC before and after therapy. DESIGN: Forty-one patients with UC from GEMINI I and LTS were studied before and at three time points (weeks 6/12/52) following VDZ therapy. Colonic biopsies were scored using the Geboes index and correlated with Mayo endoscopic subscore. Gene expression was analysed using Affymetrix gene arrays. RESULTS: Fifty-five per cent of patients achieving endoscopic healing (= Mayo endoscopic subscore 0-1) with VDZ at the studied time points also had histological healing (= Geboes grade 0-1). In most healers, some residual histological changes (eg, disturbed architecture and increased mononuclear cell infiltrate) were still observed, although this was less at week 52. VDZ restored expression of many inflammatory genes in patients with endoscopic healing only at week 52 and not before. In VDZ healers, the expression of many genes remained dysregulated at weeks 6/12/52 compared with controls. CONCLUSIONS: VDZ induces histological healing in >50% of patients with endoscopic healing, with maximal effect at week 52. VDZ also restored, although incompletely, the colonic expression of many immune-related genes in patients with UC achieving endoscopic healing at week 52. However, persistent histological and gene dysregulations did remain even in healers, suggesting that maintenance therapy will be necessary to control the intestinal inflammation. TRIAL REGISTRATION NUMBERS: NCT00783718 and NCT00790933; post-results.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Colite Ulcerativa/tratamento farmacológico , Fármacos Gastrointestinais/uso terapêutico , Mucosa Intestinal/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Adulto , Idoso , Anticorpos Monoclonais Humanizados/farmacologia , Biópsia , Colite Ulcerativa/metabolismo , Colite Ulcerativa/patologia , Colo/patologia , Colonoscopia , Método Duplo-Cego , Feminino , Fármacos Gastrointestinais/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Estudo de Associação Genômica Ampla/métodos , Humanos , Infliximab/uso terapêutico , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Mucosa Intestinal/fisiologia , Masculino , Pessoa de Meia-Idade , Análise de Componente Principal , Resultado do TratamentoRESUMO
BACKGROUND: Intestinal barrier defects are common in patients with inflammatory bowel disease (IBD). To identify which components could underlie these changes, we performed an in-depth analysis of epithelial barrier genes in IBD. METHODS: A set of 128 intestinal barrier genes was selected. Polygenic risk scores were generated based on selected barrier gene variants that were associated with Crohn's disease (CD) or ulcerative colitis (UC) in our study. Gene expression was analyzed using microarray and quantitative reverse transcription polymerase chain reaction. Influence of barrier gene variants on expression was studied by cis-expression quantitative trait loci mapping and comparing patients with low- and high-risk scores. RESULTS: Barrier risk scores were significantly higher in patients with IBD than controls. At single-gene level, the associated barrier single-nucleotide polymorphisms were most significantly enriched in PTGER4 for CD and HNF4A for UC. As a group, the regulating proteins were most enriched for CD and UC. Expression analysis showed that many epithelial barrier genes were significantly dysregulated in active CD and UC, with overrepresentation of mucus layer genes. In uninflamed CD ileum and IBD colon, most barrier gene levels restored to normal, except for MUC1 and MUC4 that remained persistently increased compared with controls. Expression levels did not depend on cis-regulatory variants nor combined genetic risk. CONCLUSIONS: We found genetic and transcriptomic dysregulations of key epithelial barrier genes and components in IBD. Of these, we believe that mucus genes, in particular MUC1 and MUC4, play an essential role in the pathogenesis of IBD and could represent interesting targets for treatment.
Assuntos
Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/patologia , Mucosa Intestinal/patologia , Transcriptoma , Adulto , Idoso , Bélgica , Estudos de Casos e Controles , Colo/patologia , Feminino , Humanos , Íleo/patologia , Modelos Logísticos , Masculino , Análise em Microsséries , Pessoa de Meia-Idade , Mucina-1/genética , Mucina-4/genética , Polimorfismo de Nucleotídeo ÚnicoRESUMO
The specific phenotype of mature differentiated beta cells not only depends on the specific presence of genes that allow beta cell function but also on the selective absence of housekeeping genes ("disallowed genes") that would interfere with this function. Recent studies have shown that both histone modifications and DNA methylation via the de novo methyltransferase DNMT3A are involved in repression of disallowed genes in neonatal beta cells when these cells acquire their mature phenotype. It is unknown, however, if the environmental influence of advanced age, pregnancy and the metabolic stress of high fat diet or diabetes could alter the repression of disallowed genes in beta cells. In the present study, we show that islet disallowed genes-which are also deeply repressed in FACS-purified beta cells-remain deeply repressed in animals of advanced age and in pregnant females. Moreover, the stability of this repression was correlated with strong and stable histone repression marks that persisted in islets isolated from 2 year old mice and with overall high expression of Dnmt3a in islets. Furthermore, repression of disallowed genes was unaffected by the metabolic stress of high fat diet. However, repression of about half of the disallowed genes was weakened in 16 week-old diabetic db/db mice. In conclusion, we show that the disallowed status of islet genes is stable under physiological challenging conditions (advanced age, pregnancy, high fat diet) but partially lost in islets from diabetic animals.
Assuntos
Envelhecimento/fisiologia , Metilação de DNA/genética , Diabetes Mellitus/metabolismo , Dieta Hiperlipídica , Código das Histonas/genética , Células Secretoras de Insulina/metabolismo , Estresse Fisiológico/fisiologia , Animais , DNA (Citosina-5-)-Metiltransferases/metabolismo , DNA Metiltransferase 3A , Feminino , Teste de Tolerância a Glucose , Insulina/metabolismo , Células Secretoras de Insulina/citologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , GravidezRESUMO
Steviol glycosides (SGs), such as stevioside and rebaudioside A, are natural, non-caloric sweet-tasting organic molecules, present in extracts of the scrub plant Stevia rebaudiana, which are widely used as sweeteners in consumer foods and beverages. TRPM5 is a Ca2+-activated cation channel expressed in type II taste receptor cells and pancreatic ß-cells. Here we show that stevioside, rebaudioside A and their aglycon steviol potentiate the activity of TRPM5. We find that SGs potentiate perception of bitter, sweet and umami taste, and enhance glucose-induced insulin secretion in a Trpm5-dependent manner. Daily consumption of stevioside prevents development of high-fat-diet-induced diabetic hyperglycaemia in wild-type mice, but not in Trpm5-/- mice. These results elucidate a molecular mechanism of action of SGs and identify TRPM5 as a potential target to prevent and treat type 2 diabetes.
Assuntos
Diterpenos do Tipo Caurano/farmacologia , Glucosídeos/farmacologia , Células Secretoras de Insulina/efeitos dos fármacos , Edulcorantes/farmacologia , Canais de Cátion TRPM/efeitos dos fármacos , Paladar/efeitos dos fármacos , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Dieta Hiperlipídica , Feminino , Células HEK293 , Humanos , Insulina/metabolismo , Secreção de Insulina , Células Secretoras de Insulina/metabolismo , Masculino , Camundongos , Camundongos Knockout , Técnicas de Patch-Clamp , Canais de Cátion TRPM/metabolismoRESUMO
AIMS/HYPOTHESIS: Type 1 diabetes is an endocrine disease where a long preclinical phase, characterised by immune cell infiltration in the islets of Langerhans, precedes elevated blood glucose levels and disease onset. Although several studies have investigated the role of the immune system in this process of insulitis, the importance of the beta cells themselves in the initiation of type 1 diabetes is less well understood. The aim of this study was to investigate intrinsic differences present in the islets from diabetes-prone NOD mice before the onset of insulitis. METHODS: The islet transcriptome and proteome of 2-3-week-old mice was investigated by microarray and 2-dimensional difference gel electrophoresis (2D-DIGE), respectively. Subsequent analyses using sophisticated pathway analysis and ranking of differentially expressed genes and proteins based on their relevance in type 1 diabetes were performed. RESULTS: In the preinsulitic period, alterations in general pathways related to metabolism and cell communication were already present. Additionally, our analyses pointed to an important role for post-translational modifications (PTMs), especially citrullination by PAD2 and protein misfolding due to low expression levels of protein disulphide isomerases (PDIA3, 4 and 6), as causative mechanisms that induce beta cell stress and potential auto-antigen generation. CONCLUSIONS/INTERPRETATION: We conclude that the pancreatic islets, irrespective of immune differences, may contribute to the initiation of the autoimmune process. DATA AVAILABILITY: All microarray data are available in the ArrayExpress database ( www.ebi.ac.uk/arrayexpress ) under accession number E-MTAB-5264.
Assuntos
Ilhotas Pancreáticas/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Estado Pré-Diabético/metabolismo , Proteômica/métodos , Animais , Hidrolases/genética , Hidrolases/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Modelos Teóricos , Estado Pré-Diabético/patologia , Desiminases de Arginina em Proteínas , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Eletroforese em Gel Diferencial BidimensionalRESUMO
BACKGROUND AND AIMS: Increased levels of tissue inhibitor of metalloproteinase-1 [TIMP-1] have been detected in both inflammatory and fibrotic lesions in Crohn's disease. In a murine model of chronic inflammation, fibrosis was associated with an increase in TIMP-1 and inhibition of matrix metalloproteinase [MMP]-mediated degradation. We investigated the effect of TIMP-1 deficiency in acute and chronic murine models of colitis. METHODS: Colitis was induced via oral administration of dextran sodium sulphate [DSS] to B6.129S4-Timp1tm1Pds/J knock-out [KO] and C57BL/6J wild-type [WT] mice. Levels of inflammation and fibrosis were assessed and gelatin zymographies and gene expression microarrays were performed. RESULTS: Compared with WT mice, TIMP-1 KO mice had higher inflammatory parameters after acute DSS administration and developed less fibrosis after chronic DSS administration. MMP-2 levels were increased in WT versus TIMP-1 KO mice with acute colitis, whereas a trend for higher proMMP-9 levels was observed in WT versus TIMP-1 KO mice with chronic colitis. In control conditions, several immune-related genes [e.g Ido1, Cldn8] were differentially expressed between young TIMP-1 KO and WT mice, but to a lesser extent between older TIMP-1 KO and WT mice. In response to DSS, the gene expression pattern was significantly different between young TIMP-1 KO and WT mice, whereas it was similar in older TIMP-1 KO and WT mice. CONCLUSIONS: TIMP-1 deficiency leads to differential expression of immune-related genes and to attenuated development of fibrosis. Unravelling the role of TIMP-1 in intestinal remodelling is necessary to develop more effective and more targeted therapeutic strategies for intestinal fibrosis.